Introduction to Molecular Biology: From Cells to Atoms - Anthony Rees, Michael Sternberg 2002

Nucleic Acids and Genes
Translation of the Genetic Code

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Fig. 25.1.

We will describe The Mechanism of Translation in prokaryotes using E. coli as an example.

The starting signal for Protein Synthesis is the AUG codon on the mRNA, which encodes Methionine (Met) [sometimes this is the GUC codon for valine (Val)]. In the growing polypeptide chain, the first amino acid residue is always either Met or Val. This naturally raises the question: how does The Cell distinguish the start signal from AUG or GUC codons located within the middle of the mRNA molecule? This problem is solved using a modified form of Met (or Val) and a special initiator tRNA (see below).

Formylmethionine (fMet) is the modified form of Met with which protein synthesis begins. It attaches to specific types of tRNA molecules (tRNA_f), which differ from tRNA_Met molecules used to incorporate Met into the inner part of the polypeptide chain. Both tRNA_f and tRNA_Met recognize the AUG codon, but only tRNA_f is capable of binding to the start codon AUG.

Initiation of protein synthesis begins with The formation of the initiation complex on the 30S subunit, consisting of mRNA, the 30S ribosomal subunit, and the aminoacyl-tRNA_f molecule with attached fMet, which binds to the P site. The next step is the joining of the 50S subunit, resulting in the Formation of the 70S initiation complex. The energy source for the initiation of protein synthesis is the Hydrolysis of GTP to GDP and P_i. Several additional Proteins, called initiation factors (IF1, IF2, and IF3), are required at this stage. Elongation is the sequential Incorporation of Amino acid residues into the growing polypeptide chain. Each elongation step consists of three phases: 1) codon recognition, 2) peptide bond formation, and 3) translocation (Fig. 25.1).

Codon recognition involves the binding of the anticodon of the incoming aminoacyl-tRNA molecule to the codon in the unoccupied A site on the ribosome. To attach to the ribosome, the tRNA with its attached amino acid must first form a complex with a protein called elongation factor EF-Tu or EF1, which must be pre-activated by GTP. After the entire tRNA-EF1 ∙ GTP complex binds to the A site of the ribosome, GTP is hydrolyzed to GDP and P_i, supplying the energy required for this stage of elongation. The EF1 ∙ GDP factor, no longer able to bind to tRNA, leaves the ribosome, leaving the aminoacyl-tRNA behind. Regeneration of the activated factor EF1 is catalyzed by a second elongation factor, EF-Ts or EF2, which replaces GDP in the inactive complex, resulting in the formation of the EF1 ∙ EF2 complex. A diagram illustrating this sequence of events is presented in Fig. 25.2.

Fig. 25.2.

Peptide bond formation occurs only when both the A and P sites are occupied by aminoacyl-tRNA molecules. Part of the 50S subunit Functions as the enzyme peptidyl transferase, which catalyzes peptide bond formation According to the scheme shown in Fig. 25.3. As a result of this reaction, the growing polypeptide chain becomes attached to the tRNA at the A site, while the tRNA at the P site is released from the complex with the peptide and carries an —OH group at its 3'-end (Fig. 25.3).

Translocation involves three steps catalyzed by another elongation factor, EF-G (EF3), and energetically coupled to GTP hydrolysis. First, the uncharged tRNA at the P site leaves the ribosome, then the peptidyl-tRNA molecule moves from the A site to the P site, and finally, the ribosome shifts along the mRNA by three nucleotide residues toward the 3'-end. These three steps free up the A site and expose the next codon, allowing the next elongation cycle to begin.

Termination, or the completion of synthesis, is triggered by the UAA, UGA, or UAG codons. Nature does not provide tRNA molecules with anticodons corresponding to these codons. Instead of chain elongation continuing, termination is catalyzed by special proteins called termination factors, which recognize the stop codons when the A site is vacant. These factors alter the Specificity of the peptidyl transferase enzyme so that the bond between the terminal peptide and the tRNA is hydrolyzed, and the released polypeptide chain diffuses away from the ribosome. This is followed by the dissociation of the mRNA-ribosome complex. Next, the ribosome dissociates into 30S and 50S subunits. Upon reassociation of these subunits with another mRNA molecule, the entire protein synthesis cycle begins anew. EUKARYOTIC TRANSLATION, which takes place in the Cytoplasm, involves the same stages as prokaryotic translation. The main difference here is that the first residue in the growing polypeptide chain is Met rather than fMet. Nevertheless, There are also Two Types of tRNA molecules that recognize the AUG codon: one when the codon is an initiator, and another when it encodes Met to be incorporated within the middle of the growing polypeptide chain. Different proteins act as initiation and elongation factors. Another significant difference is that Ribosomes are larger (80S) in the eukaryotic cytoplasm.

Fig. 25.3.

In Mitochondria and METABOLISM/14.html">Chloroplasts, translation occurs within the Organelles themselves. The ribosomes they contain are 70S particles and resemble bacterial ribosomes. Initiation utilizes fMet.